Department of Chemistry, Physical &Theoretical Chemistry Laboratory, University of Oxford, South Parks Road, Oxford, Oxfordshire OX1 3QZ, UK.
Department of Chemistry-BMC, Uppsala University, Box 576, Uppsala SE-751 23, Sweden.
Nat Commun. 2017 Jan 10;8:13993. doi: 10.1038/ncomms13993.
Na/H antiporters are found in all kingdoms of life and exhibit catalysis rates that are among the fastest of all known secondary-active transporters. Here we combine ion mobility mass spectrometry and molecular dynamics simulations to study the conformational stability and lipid-binding properties of the Na/H exchanger NapA from Thermus thermophilus and compare this to the prototypical antiporter NhaA from Escherichia coli and the human homologue NHA2. We find that NapA and NHA2, but not NhaA, form stable dimers and do not selectively retain membrane lipids. By comparing wild-type NapA with engineered variants, we show that the unfolding of the protein in the gas phase involves the disruption of inter-domain contacts. Lipids around the domain interface protect the native fold in the gas phase by mediating contacts between the mobile protein segments. We speculate that elevator-type antiporters such as NapA, and likely NHA2, use a subset of annular lipids as structural support to facilitate large-scale conformational changes within the membrane.
Na+/H+ 反向转运蛋白存在于所有生命领域，其催化速率在所有已知的次级主动转运蛋白中属于最快的之列。在这里，我们结合离子淌度质谱和分子动力学模拟来研究来自嗜热栖热菌的 Na+/H+ 交换蛋白 NapA 的构象稳定性和脂质结合特性，并将其与原型反向转运蛋白 NhaA（来自大肠杆菌）和人类同源物 NHA2 进行比较。我们发现，NapA 和 NHA2（但不是 NhaA）形成稳定的二聚体，并且不选择性地保留膜脂质。通过将野生型 NapA 与工程变体进行比较，我们表明，蛋白质在气相中的展开涉及到结构域间接触的破坏。结构域界面周围的脂质通过介导可移动蛋白片段之间的接触，在气相中保护天然折叠。我们推测，像 NapA 这样的提升式反向转运蛋白，以及可能的 NHA2，利用一组环形脂质作为结构支撑，以促进膜内的大规模构象变化。
